The role of Ca2+, PO43- and F- in nano-crystal formation in the kidney
The groundwater consumed in the affected communities has high dissolved solids and electrical conductivity, Ca2+, phosphate (PO43), and fluoride (F−). For example, median range of these in groundwater, in affected regions are, TDS 500 mg/L, EC 1,800 µS/cm, calcium 75 mg/L, Mg2+ 50 mg/L, alkalinity 250 mg/L, SO42− 200 mg/L, F− 0.8 mg/L, and PO43− 0.025 mg/L.8 Most are unhealthy to consume chronically and are above the EPA and WHO mandated safety standards.8 Besides, because of the unpalatability of water, people in these harsh/hot tropical climatic regions consume less water, but daily alcohol consumption makes them chronically dehydrated.
Conditions in renal tissues are conducive to calcium phosphate (CaPO4) nano-mineral formation. Although CaPO4-hydroxyapatite formation in kidneys is not unusual: these structures are unstable [Ca5(PO4)3(OH) = Ca2+ + 3PO43− + OH−]. Groundwater in these regions also contains higher concentrations of F−but is sporadic. At the right conditions, F− can substitute hydroxyl or PO4 groups in CaPO4 hydroxyapatite to form more stable fluorapatite crystals through ionic and hydrogen bonds [fluoroapatite: 5Ca2+ + 3PO43− + F−Ca5(PO4)3−F].
This phenomenon is analogous to incorporating F− into CaPO4 apatite in the enamel component in teeth and skeletal tissues. Despite requiring a minute concentration of F-, it produces stable fluoroapatite crystals resistant to decay and strengthens existing hydroxyapatite scaffolds. Consequent cell-mediated fluorapatite, carbonatoapatite, and other nano-crystals make these tissues stiffer.9 Nevertheless, excess incorporation of F− causes brownish discolouration of teeth and brittleness of bones.10
Nano-crystal and nano-tube formation in renal tissues
Ultra-structural studies in human kidney tissues with end-stage CKD consistently reported 50 to 1,500 nm multi-lamellar mineral particles.11 Presence of CaPO4 crystals is also reported in renal biopsy samples in those with several chronic renal diseases.7,12−14 These nano-mineral particles of polycrystalline CaPO4 are produced in extracellular matrices and tubular cells.13,14 Fig. 1 is a conceptual diagram illustrating Geo-Bio interactions, pathways, and conditions necessary to precipitate CaPO4 nano-crystal in renal tissues in renal tubules, causing CKD-CTN.
Illustrates the Geological and Biological (Geo-Bio) pathways of forming nano-tubes and nano-crystals in CKD-CTN. A conceptual diagram illustrating pathways for the formation of calcium phosphate (CaPO4) nano-crystals in kidney tubules causing CKD-CTN: (a) Interaction of rocks containing minerals rich in F−, Ca2+, Mg2+, PO43− with groundwater; (b) Long duration of water-rock interaction; (c) Consumption of water with high concentrations of ions; (D) Biochemical reactions and super-saturation, precipitating CaPO4 nano-crystals and nano-tubes in kidney tubules; (e) Factors enhancing the nano-mineral precipitation; (f) Low-grade chronic inflammation and tissue fibrosis; and (g) Onset of chronic renal failure (from Wimalawansa & Dissanayake, 2022; Frontiers of Water and human health).
Growth of hydroxy- and fluorapatite nanoparticles occurs slowly in vivo. These are inflammatogenic and thus, attract fibroblast and leukocytes, causing low-grade inflammation, tissue fibrosis, and eventually renal failure. Once nano-crystal complexes are precipitated in vivo, these attract matrix proteins 15 and incorporate these into hydroxy-fluorapatite, further strengthening nano-minerals that become resistant to enzymatic degradation, thus preventing their dissolution.16,17 These allow fluoroapatite crystals to grow, eventually blocking and perforating renal tubules, causing cellular apoptosis.
Mechanisms of formation of nano-tubes in renal tubules
In 2019 the authors postulated that repeated chronic dehydration generated peaks of Ca2+ and (PO4)-3, which initiates the formation of nano-crystals nano-tubes and their growth. This chronic inflammatory process gradually produces tissue fibrosis, causing CKD-CTN and premature deaths from renal failure-associated complications.4 Water consumption with higher mineral content for more than a decade leads to increased tissue, intra-tubular Ca2+, and other minerals increasing the risks for mineral crystal formation in kidneys. Anecdotally, the provision of potable water through reverse osmosis in several affected villages led to over a 50% reduction in the incidence of CKDmfo within two years.18
In CKD-CTN-prone dry zonal regions in tropical countries, groundwater has significantly higher total dissolved substances (TDS) and minerals, conducive to forming CaPO4 hydroxyapatite minerals.4 In addition, those who live in the CKD-CTN-affected villagers have a higher intestinal fractional absorption of minerals secondary to malnutrition. As the Ca2+ PO43- product exceeds a solubility threshold at the body temperature, the formation of CaPO4 nano-particles occurs. Besides CaPO4, super-saturation with CaOx, urates, or other minerals in renal tubules and interstitial tissues, initiates and propagates complexed nano-minerals in renal tubules.
Chronic dehydration continues due to lesser water consumption because of unpalatability of hard water, excessive sweating, and alcohol abuse leads to the formation of concentrated urine and reduction of the flow (stagnation) in renal tubules, increasing the likelihood of formation of nano-minerals. It takes more than a decade to develop a symptomatic renal disease and full-blown renal failure.19,20 When nano-crystals exceed 1 mm, such accumulations can become visible with in vivo imaging methods, such as high-resolution computed tomography.7,12,21 However, nanoparticles are too small to be visible in routine radiological investigations.
Magnesium deficiency increases the risks of renal nano-crystal formation
Mg2+ competes with Ca2+ in many physiological situations and is a cofactor for most enzymatic reactions and the release of hormones from endocrine cells. Hypomagnesemia enhances the accumulation of Ca2+ within renal cells. Moreover, the lack of antagonist effect of Mg2+on Ca2+ increases the risk of CaPO4 nano-crystal formation in soft tissues, particularly in the kidneys, due to the super-saturation of stagnant fluid. In addition, Mg2+ antagonise the formation of calcium and nano-crystal in kidneys.
When rats fed an Mg-deficient diet, the harmful effects of calcium are accentuated and worsened by low dietary potassium. Likewise, as an adverse effect, circulatory Mg2+ concentration is reduced when rats were fed with high calcium diets.22 In contrast, renal accumulation of calcium (CaPO4 and precipitation of other minerals) is mitigated with Mg2+ supplements. In contrast, the intracellular magnesium concentration is necessary for physiological activities and lower circulatory Mg2+ concentrations that reflect low Mg2+ levels in tissues that facilitate CaPO4 precipitation. In addition to dietary factors, these data emphasise the importance of the quality of drinking water, multiple ion interactions and their ratios and competition in vivo physiological situations. These either mitigate or aggravate the ionic reactions and clinical outcomes.
Dietetic fructose in renal tubular CaPO4 nanotube formation:
Rats fed on an Mg-deficient diet experienced hypomagnesemia, reduced renal tissue Mg2+, and increased Ca2+ concentrations.23 This was worsened when rats were fed with glucose or fructose compared to those fed on unrefined, complex carbohydrates. In the fructose-fed group, renal Ca content was eight times greater than in the control group.23 High fructose diet also worsens existing hypercalcemia, hypercalciuria, and hypomagnesuria, enhancing CaPO4 crystallisation in kidneys.
Excess fructose in the presence of low Mg2+ increases risks for nephrocalcinosis and the likelihood of renal failure. Higher concentrations of F- increase Ca2+ precipitation in soft tissues, arteries, and kidneys. Those who live in regions with a higher prevalence of CKD-CTN are economically poor, and their diets are predominantly carbohydrates. In addition, these diets are deficient in micronutrients and Mg2; fructose is a significant component of carbohydrates. In the presence of low cell/tissue Mg2+ concentration, higher fructokinase activity further increases the risks of renal calcification.5 For mineral precipitation, it is necessary to have prolonged dehydration and low urine output. Urine pH of around 6.8, hypercalcemia, hypercalciuria, hypomagnesuria, and low urinary Mg2+-to-Ca2+ ratio, are favourable for CaPO4 precipitation in the kidney.24
Synergistic ion interactions exacerbate the nano-crystal formation
Physiological concentrations of Mg2+ counteract the adverse effects of increased intracellular Ca2+ concentration and modulation of Ca2+ channels. Hypomagnesemia increases N-methyl-D-aspartate receptors and nuclear factor-kappaB activity, stimulating the renin-angiotensin system and reducing renal blood flow.25 Aggregated apatite nano-tubes attach to the luminal cell surfaces,13 causing blockage and/or rupturing renal tubules and reducing renal functions.
Insufficiency of crystallisation modulators, such as Tamm-Horsfall protein, osteopontin, sodium phosphate co-transporter, or sodium-hydrogen exchanger regulatory factor-1, also increases the risks of tubular and interstitial nephrocalcinosis.26 Whereas the crystal-induced interstitial tissue inflammation and oxidative stress further reduce intra-renal blood flow via activation of the renin-angiotensin system, causing further impairment of renal functions.
Other contributory factors
Renal failure increases PO4 retention, which leads to increased Klotho levels. Meanwhile, advanced renal failure disrupts fibroblast growth factor-23 (FGF23) signalling that increases the accumulation of PO4.27 Therefore, modulation of Klotho activity should be investigated as a target for intervention in those with moderate CKD-CTN. For example, using Klotho or its synthetic agonist reduces PO4 toxicity and severity in CKD-CTN and accelerated ageing.28 Hypomagnesemia worsens retention and the Klotho-induced PO4 tubular load and renal functions. Therefore, prophylactically correcting Mg2+ deficiency could also be a cost-effective approach for those living in endemic areas to reduce PO4 load and an economical way to prevent and reverse CKD-CTN in its early stages. Figure 2 illustrates the fundamental mechanisms and pathways involved in forming CaPO4 crystals in renal tissue.
It is not climate change but the associated disasters that arose from willful destruction of the environment through uncontrolled development and agriculture and irresponsible behaviour of people that endanger nature, causing many preventable human diseases. Since CKD-CTN originates from an environmental cause—natural Geo-Bio interactions (Figure 1)—it is feasible to reduce the risk by addressing the root cause: without embarking on expensive pharmaceutical agents. As we and others have previously reported,1,29,30 The most critical intervention is supplying potable water to all affected regions.20,31 This can be achieved via mentioned cost-benefits analysis and practical steps to help eradicate CKD-CTN,32,20 and adopting previously described economic longer-term chronic diseases management programs.32,33 The following section describes ways to control and eradicate CKD-CTN.
Eradication of CKD-CTN is straightforward
Interventions for CKD-CTN in affected regions need to focus on providing potable water at an affordable price and educating them on ways to avoid chronic dehydration. Providing centrally purified pipe-borne water to affected regions is expensive and estimated to take more than three decades. In the interim, the most cost-effective and practical way to provide a scalable potable water supply is through reverse osmosis or water descaling/softening plants to remove hardness, also capable of removing excess F-, scattered across affected regions.34-36 A program to educate the public, starting from school and extending to the community, on avoiding harmful behaviour like daily alcohol intake is critical to preventing the younger generation from getting affected.
In addition, when a breadwinner or any family member acquires CKD-CTN, their expenses escalate, and their income dwindles. This initiates a vicious cycle within the family that affects children’s education and ability to do manual activities and generate income. Consequently, it increases the poverty that escalates malnutrition: this is a universal phenomenon in all affected regions.37,38 Therefore, programs to alleviate CKD-CTN must encompass poverty alleviation. The authors recommend implementing straightforward, cost-effective muti-prong programs to prevent this fatal CKD-CTN,4,31,38,39 of the previously described.39 A broader holistic and affirmative approach based on the concepts described,20,36 encompassing education, awareness, prevention of environmental pollution, lessening malnutrition, correcting unhealthy behaviours and habits acquired during the past four decades, and providing clean water would be sufficient to reduce the incidence and eradicating this deadly disease rapidly.40-42